The upfield shift (2.6 ppm) observed upon formation of the ternary complex: L. casei dihydrofolate reductase-TPN-folic acid-13C-benzoyl carbonyl is due to the ring current anisotropy of Phe-49, as revealed by the crystal structure of L. casei dihydrofolate reductase-TPNH-MTX. Aminopterin with the same 13C label shows a shift of 3.0 ppm upfield for the ternary complex in solution, indicating a similar geometry in the solution structure of this ternary complex. Formation of the respective ternary complexes from 3', 5',-dibromofolic acid-13C-benzoyl carbonyl and 3', 5',-dibromoaminopterin-13C-benzoyl carbonyl should interfere with the packing of Phe-30, a conserved residue in all known dihydrofolate reductases, and may reveal differences in solution structure. The dibromo compounds also serve as probes of Trp-21 which will be studied by completion of the preparation by fermentation of L. casei dihydrofolate reductase containing 13C in the 3' position of its four tryptophane residues. The preparation of folic acid and Aminopterin labeled at the 4 position with 13C will reveal possible differences in bonding of the 2-amino-4-hydroxy compounds and the 2, 4-diamino inhibitors of this enzyme. The 13C label at the 2 position of these tow compounds will monitor the bonding of Asp-26 at the N1-N2 region, and may reveal differences in solution structure.